In order to develop T-cell therapies against non-melanoma cancers, such as ovarian, breast, and colon cancer, we have developed chimeric antibody/T- cell receptor genes which combine variable regions from mAb with T-cell signaling chains. Previously, we demonstrated that T-cells transduced with chimeric receptor genes are redirected to recognize new targets. Over the past year, we have further defined the conditions necessary to transduce primary peripheral blood lymphocytes, and have taken steps towards applying this technology to human cancer patients. A clinical protocol utilizing this strategy for ovarian cancer patients has so far been approved by the NIH Biosafety Committee, Institutional Review Board, and the NIH Recombinant Advisory Committee. Retroviral transduction of primary lymphocytes has been improved by utilizing retroviral vectors that contain internal ribosomal entry sites (IRES) instead of traditional vectors containing 2 distinct promoters. In addition, we have studied the relative expression of the gibbon ape leukemia virus (GALV) and amphotropic retroviral receptors in T-cells, and have determined that the GALV receptor is expressed 30-fold higher than the amphotropic receptor and that utilizing the GALV envelope results in enhanced transduction efficiencies in lymphocytes. We have also studied mice reconstituted with bone marrow cells transduced with chimeric T-cell receptor genes. We have so far demonstrated that activated T-cells from the spleens of these mice function against the appropriate antigen, defined by the chimeric T-cell receptor. Future studies will focus on the function of transduced monocytes and neutrophils from these mice. We have continued studies to modify lymphocytes with growth factor receptor genes in order to support them in vivo with ligands that are less toxic than IL-2. So far, we have demonstrated that IL-2 dependent T-cell lines transduced with either the GM-CSF receptor or chimeric erythropoietin (EPO)/IL-2 receptor gene can grow in response to GM-CSF or EPO, respectively, in the absence of IL-2. Future studies will analyze the in vivo survival of these T-cells using the appropriate alternative ligands.